Two major metabolites in man of the widely used antituberculosis drug, isoniazid, are monoacetylhydrazine and diacetylhydrazine. Because either one or both of these hydrazines may be responsible for isoniazid liver injury, we are developing procedures to monitor these two metabolites in human plasma and urine. Gas chromatographic assays have been developed for both monoacetylhydrazine and diacetylhydrazine with sensitivities for plasma assays ranging down to 100 ng/ml. The assay for diacetylhydrazine is erratic, however, and a more specific and accurate assay for both mono- and diacetylhydrazine is being developed using stable isotope internal standards and combined gas chromatography-mass spectrometry. Once conditions are worked out we will monitor the effects of inducers and inhibitors of drug metabolism on the disposition in vivo of these potentially toxic metabolites. Such studies offer the possibility that therapeutic regimens can be developed which decrease either the production of such metabolites or their subsequent biotransformation to reactive metabolites.